Binding of terbium and of an elastase inhibitor to bovine pancreatic subunit III, an inactive protease E.
Using the Tb3+ luminescence technique, we showed that bovine subunit III, a defective pancreatic serine endopeptidase-like protease, possessed a single metal ion binding site able to bind Tb3+ with a high affinity comparable to that of porcine elastase. The topology of the metal ion binding site in subunit III is predicted from sequence homologies and modeling experiments based on the known crystallographic three-dimensional structures of the equivalent sites in porcine elastase 1 and bovine beta-trypsin. Moreover, the Tb3+ luminescence technique in parallel to a 19F NMR investigation, allowed us to measure the binding of a very potent specific inhibitor of porcine elastase (trifluoroacetyl-L-lysyl-alanyl p-trifluoromethylphenylanilide) to bovine subunit III. These results confirm that, although devoid of any specific activity, subunit III might possess a conformation close to that of an active enzyme and further support the analogy between subunit III and an elastase-like family.
Chapus
C
C
Centre de Biochimie et de Biologie Moléculaire du Centre National de la Recherche Scientifique, Marseille, France.
Kerfelec
B
B
Dimicoli
J L
JL
D000595:Amino Acid Sequence; D000813:Anilides / Q000378:metabolism*; D000818:Animals; D001665:Binding Sites; D002417:Cattle; D006801:Humans; D007700:Kinetics; D046911:Macromolecular Substances; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D010179:Pancreas / Q000201:enzymology*; D010196:Pancreatic Elastase / Q000037:antagonists & inhibitors; D011485:Protein Binding; D012689:Sequence Homology, Nucleic Acid; D012697:Serine Endopeptidases / Q000302:isolation & purification / Q000378:metabolism*; D015842:Serine Proteinase Inhibitors; D013050:Spectrometry, Fluorescence; D013725:Terbium / Q000378:metabolism*
D000813:Anilides; D046911:Macromolecular Substances; D015842:Serine Proteinase Inhibitors; D013725:Terbium; C037928:trifluoroacetyllysylalanine-trifluoromethylphenylanilide; D012697:Serine Endopeptidases; C028734:protease E; D010196:Pancreatic Elastase